Pheromones induce sexually dimorphic neuroendocrine responses, such as LH secretion. However, the neuronal network by which pheromones are converted into signals that will initiate and modulate endocrine ... [more ▼]

Pheromones induce sexually dimorphic neuroendocrine responses, such as LH secretion. However, the neuronal network by which pheromones are converted into signals that will initiate and modulate endocrine changes remains unclear. We asked whether two sexually dimorphic populations in the anteroventral periventricular and periventricular nuclei (AVPV/PeN) that express kisspeptin and tyrosine hydroxylase (TH) are potential candidates that will transduce the olfactory signal to the neuroendocrine system. Furthermore, we assessed whether this transduction is sensitive to perinatal actions of estradiol by using female mice deficient in alpha-fetoprotein (AfpKO), which lack the protective actions of Afp against maternal estradiol. Wild-type (WT) and AfpKO male and female mice were exposed to same- versus opposite-sex odors and the expression of c-Fos was analyzed along the olfactory projection pathways as well as whether kisspeptin, TH and gonadotropin-releasing hormone (GnRH) neurons are responsive to opposite-sex odors. Male odors induced a female-typical Fos expression in target forebrain sites of olfactory inputs involved in reproduction in WT, but not in AfpKO females, whereas female odors induced a male-typical Fos expression in males of both genotypes. In WT females, opposite-sex odors induced Fos in kisspeptin and TH neurons, whereas in AfpKO females and WT males, only a lower, but still significant, Fos expression was observed in TH, but not in kisspeptin neurons. Finally, opposite-sex odors did not induce any significant Fos expression in GnRH neurons of both sexes or genotypes. Our results strongly suggest a role for fetal estrogen in the sexual differentiation of neural responses to sex-related olfactory cues. [less ▲]

The sexual differentiation of the brain is primarily driven by gonadal hormones during fetal development. Leading theories on the etiology of gender dysphoria (GD) involve deviations herein. To examine ... [more ▼]

The sexual differentiation of the brain is primarily driven by gonadal hormones during fetal development. Leading theories on the etiology of gender dysphoria (GD) involve deviations herein. To examine whether there are signs of a sex-atypical brain development in GD, we quantified regional neural gray matter (GM) volumes in 55 female-to-male and 38 male-to-female adolescents, 44 boys and 52 girls without GD and applied both univariate and multivariate analyses. In girls, more GM volume was observed in the left superior medial frontal cortex, while boys had more volume in the bilateral superior posterior hemispheres of the cerebellum and the hypothalamus. Regarding the GD groups, at whole-brain level they differed only from individuals sharing their gender identity but not from their natal sex. Accordingly, using multivariate pattern recognition analyses, the GD groups could more accurately be automatically discriminated from individuals sharing their gender identity than those sharing their natal sex based on spatially distributed GM patterns. However, region of interest analyses indicated less GM volume in the right cerebellum and more volume in the medial frontal cortex in female-to-males in comparison to girls without GD, while male-to-females had less volume in the bilateral cerebellum and hypothalamus than natal boys. Deviations from the natal sex within sexually dimorphic structures were also observed in the untreated subsamples. Our findings thus indicate that GM distribution and regional volumes in GD adolescents are largely in accordance with their respective natal sex. However, there are subtle deviations from the natal sex in sexually dimorphic structures, which can represent signs of a partial sex-atypical differentiation of the brain. [less ▲]

Sex steroid hormones act on developing neural circuits regulating the hypothalamic-pituitary-gonadal axis and are involved in hormone-sensitive behaviours. These hormones act mainly via nuclear receptors, i.e. oestrogen receptor-alpha (ERalpha) and androgen receptor (AR). By using immunohistochemistry, we analysed the expression level of ERalpha and AR throughout perinatal life [at embryonic (E) day 19 and postnatal (P) days 5-15-25] and in adulthood in several hypothalamic nuclei controlling reproduction in both wild-type (WT) and aromatase knockout (ArKO) (which cannot convert testosterone into oestradiol) mice to determine whether there are sex differences in hypothalamic ERalpha and AR expression and if so, whether these are established by oestradiol action. As early as E19, ERalpha immunoreactivity (-ir) was observed at same expression levels in both sexes in the anteroventral periventricular nucleus (AVPv), the medial preoptic area (MPOA), the bed nucleus of the stria terminalis (BnST), the ventrolateral part of the ventromedial hypothalamic nucleus (VMHvl) and the arcuate nucleus (ARC). Sex differences (female > male) in ERalpha-ir were only observed during the prepubertal period in the BnST (P5 to P25) and the MPOA (P15), but also in adulthood in these two brain regions. Sex differences in AR-ir (male > female) were observed at P5 in the AVPv and ARC, and at P25 in the MPOA and ARC as well as in adulthood in all hypothalamic regions analysed. In adulthood, gonadectomy and hormonal treatment (oestradiol or dihydrotestosterone) also strongly modulated ERalpha-ir and AR, respectively. Taken together, sex differences in ERalpha-ir and AR-ir were observed in all hypothalamic regions analysed, but most likely do not reflect oestradiol actions since ArKO mice of both sexes showed very similar expression levels as WT mice throughout perinatal development. This article is protected by copyright. All rights reserved. [less ▲]

Sex hormones, androgens in particular, are hypothesized to play a key role in the sexual differentiation of the human brain. However, possible direct effects of the sex chromosomes, that is, XX or XY ... [more ▼]

Sex hormones, androgens in particular, are hypothesized to play a key role in the sexual differentiation of the human brain. However, possible direct effects of the sex chromosomes, that is, XX or XY, have not been well studied in humans. Individuals with complete androgen insensitivity syndrome (CAIS), who have a 46,XY karyotype but a female phenotype due to a complete androgen resistance, enable us to study the separate effects of gonadal hormones versus sex chromosomes on neural sex differences. Therefore, in the present study, we compared 46,XY men (n = 30) and 46,XX women (n = 29) to 46,XY individuals with CAIS (n = 21) on a mental rotation task using functional magnetic resonance imaging. Previously reported sex differences in neural activation during mental rotation were replicated in the control groups, with control men showing more activation in the inferior parietal lobe than control women. Individuals with CAIS showed a female-like neural activation pattern in the parietal lobe, indicating feminization of the brain in CAIS. Furthermore, this first neuroimaging study in individuals with CAIS provides evidence that sex differences in regional brain function during mental rotation are most likely not directly driven by genetic sex, but rather reflect gonadal hormone exposure. [less ▲]

Click-evoked otoacoustic emissions (CEOAEs) are echo-like sounds that are produced by the inner ear in response to click-stimuli. CEOAEs generally have a higher amplitude in women compared to men and ... [more ▼]

Click-evoked otoacoustic emissions (CEOAEs) are echo-like sounds that are produced by the inner ear in response to click-stimuli. CEOAEs generally have a higher amplitude in women compared to men and neonates already show a similar sex difference in CEOAEs. Weaker responses in males are proposed to originate from elevated levels of testosterone during perinatal sexual differentiation. Therefore, CEOAEs may be used as a retrospective indicator of someone's perinatal androgen environment. Individuals diagnosed with Gender Identity Disorder (GID), according to DSM-IV-TR, are characterized by a strong identification with the other gender and discomfort about their natal sex. Although the etiology of GID is far from established, it is hypothesized that atypical levels of sex steroids during a critical period of sexual differentiation of the brain might play a role. In the present study, we compared CEOAEs in treatment-naive children and adolescents with early-onset GID (24 natal boys, 23 natal girls) and control subjects (65 boys, 62 girls). We replicated the sex difference in CEOAE response amplitude in the control group. This sex difference, however, was not present in the GID groups. Boys with GID showed stronger, more female-typical CEOAEs whereas girls with GID did not differ in emission strength compared to control girls. Based on the assumption that CEOAE amplitude can be seen as an index of relative androgen exposure, our results provide some evidence for the idea that boys with GID may have been exposed to lower amounts of androgen during early development in comparison to control boys. [less ▲]

The odorous steroid androstadienone, a putative male chemo-signal, was previously reported to evoke sex differences in hypothalamic activation in adult heterosexual men and women. In order to investigate ... [more ▼]

The odorous steroid androstadienone, a putative male chemo-signal, was previously reported to evoke sex differences in hypothalamic activation in adult heterosexual men and women. In order to investigate whether puberty modulated this sex difference in response to androstadienone, we measured the hypothalamic responsiveness to this chemo-signal in 39 pre-pubertal and 41 adolescent boys and girls by means of functional magnetic resonance imaging. We then investigated whether 36 pre-pubertal children and 38 adolescents diagnosed with gender dysphoria (GD; DSM-5) exhibited sex-atypical (in accordance with their experienced gender), rather than sex-typical (in accordance with their natal sex) hypothalamic activations during olfactory stimulation with androstadienone. We found that the sex difference in responsiveness to androstadienone was already present in pre-pubertal control children and thus likely developed during early perinatal development instead of during sexual maturation. Adolescent girls and boys with GD both responded remarkably like their experienced gender, thus sex-atypical. In contrast, pre-pubertal girls with GD showed neither a typically male nor female hypothalamic activation pattern and pre-pubertal boys with GD had hypothalamic activations in response to androstadienone that were similar to control boys, thus sex-typical. We present here a unique data set of boys and girls diagnosed with GD at two different developmental stages, showing that these children possess certain sex-atypical functional brain characteristics and may have undergone atypical sexual differentiation of the brain. [less ▲]

Behavioral testing methods are described for determining whether female mice can discriminate between volatile urinary pheromones of conspecifics of the same vs. opposite sex and/or in different endocrine ... [more ▼]

Behavioral testing methods are described for determining whether female mice can discriminate between volatile urinary pheromones of conspecifics of the same vs. opposite sex and/or in different endocrine conditions, for determining sexual partner preference, for quantifying receptive (lordosis) behavior, and for monitoring the expression of male-typical mounting behavior in female mice. [less ▲]

A brain circuit (the accessory olfactory system) that originates in the vomeronasal organ (VNO) and includes the accessory olfactory bulb (AOB) plus additional forebrain regions mediates many of the ... [more ▼]

A brain circuit (the accessory olfactory system) that originates in the vomeronasal organ (VNO) and includes the accessory olfactory bulb (AOB) plus additional forebrain regions mediates many of the effects of pheromones, typically comprised of a variety of non-volatile and volatile compounds, on aspects of social behavior. A second, parallel circuit (the main olfactory system) that originates in the main olfactory epithelium (MOE) and includes the main olfactory bulb (MOB) has also been shown to detect volatile pheromones from conspecifics. Studies are reviewed that point to specific roles of several different steroids and their water-soluble metabolites as putative pheromones. Other studies are reviewed that establish an adult, 'activational' role of circulating sex hormones along with sex differences in the detection and/or processing of non-steroidal pheromones by these two olfactory circuits. Persisting questions about the role of sex steroids in pheromonal processing are posed for future investigation. [less ▲]

In rodents, kisspeptin expressing neurons are localized in two hypothalamic brain nuclei [anteroventral periventricular nucleus/periventricular nucleus continuum (AVPv/PeN) and arcuate nucleus (ARC)] and modulated by sex steroids. By using wild-type (WT) and aromatase knockout mice (ArKO, which cannot convert testosterone into estradiol) and immunohistochemistry, we observed that WT females showed a continuous increase in kisspeptin peptide expression in the ARC across postnatal ages (P5 to P25), whereas WT males did not show any expression before P25. Kisspeptin peptide expression was also present in ArKO females but did not increase over this early postnatal period, suggesting that kisspeptin peptide expression in the ARC is organized by estradiol-dependent and -independent mechanisms. We also compared kisspeptin peptide expression between groups of adult male and female mice which were left gonadally intact or gonadectomized and treated or not with estradiol (E2) or dihydrotestosterone (DHT). In the ARC, kisspeptin peptide expression decreased after gonadectomy but was completely rescued by either E2 or DHT treatment in each sex/genotype. However, kisspeptin peptide expression was lower in ArKO compared to WT subjects. In the AVPv/PeN, ArKO females showed a male-typical kisspeptin peptide expression, and adult E2 treatment partially restored kisspeptin peptide expression. Finally, we showed that, after E2 treatment of WT and ArKO mice between either P5 and P15 or P15 and P25, AVPv/PeN kisspeptin peptide expression could be still masculinized at P5, but was feminized from P15 onwards. In conclusion, the two kisspeptin neuronal populations (AVPv/PeN versus ARC) seem to be differentially organized and activated by E2. [less ▲]

in Journal of Clinical Endocrinology and Metabolism (2012), 97(12), 2210-20

CONTEXT: The recent report that loss-of-function mutations in either the gene encoding neurokinin B (NKB) or its receptor (NK3R) produce gonadotropin deficiencies in humans strongly points to NKB as a key ... [more ▼]

CONTEXT: The recent report that loss-of-function mutations in either the gene encoding neurokinin B (NKB) or its receptor (NK3R) produce gonadotropin deficiencies in humans strongly points to NKB as a key regulator of GnRH release. OBJECTIVES: We used NKB immunohistochemistry on postmortem human brain tissue to determine: 1) whether the human NKB system in the infundibular nucleus (INF) is sexually dimorphic; 2) at what stage in development the infundibular NKB system would diverge between men and women; 3) whether this putative structural difference is reversed in male-to-female (MtF) transsexual people; and 4) whether menopause is accompanied by changes in infundibular NKB immunoreactivity. METHODS: NKB immunohistochemical staining was performed on postmortem hypothalamus material of both sexes from the infant/pubertal period into the elderly period and from MtF transsexuals. RESULTS: Quantitative analysis demonstrated that the human NKB system exhibits a robust female-dominant sexual dimorphism in the INF. During the first years after birth, both sexes displayed a moderate and equivalent level of NKB immunoreactivity in the INF. The adult features emerged progressively around puberty until adulthood, where the female-dominant sex difference appeared and continued into old age. In MtF transsexuals, a female-typical NKB immunoreactivity was observed. Finally, in postmenopausal women, there was a significant increase in NKB immunoreactivity compared to premenopausal women. CONCLUSION: Our results indicate that certain sex differences do not emerge until adulthood when activated by sex steroid hormones and the likely involvement of the human infundibular NKB system in the negative and positive feedback of estrogen on GnRH secretion. [less ▲]

The odorous steroid compound 4,16-androstadien-3-one (androstadienone), found in axillary sweat, was previously reported to evoke hypothalamic activation in heterosexual women, but not in heterosexual men ... [more ▼]

The odorous steroid compound 4,16-androstadien-3-one (androstadienone), found in axillary sweat, was previously reported to evoke hypothalamic activation in heterosexual women, but not in heterosexual men. However, subjects were exposed to the pure crystalline form of androstadienone, which raised the question whether the observed hypothalamic response is physiologically relevant. Therefore, in the present study, we asked whether sexually dimorphic hypothalamic responses could be measured when subjects were exposed to lower, more physiologically relevant concentrations of androstadienone. A total of 21 women and 16 men, all heterosexual, participated in our functional magnetic resonance imaging study (fMRI). Three different concentrations of androstadienone diluted in propylene glycol (10 mM "high," 0.1 mM "medium" and 0.001 mM "low") were delivered to the subjects' nostrils using a computer-controlled stimulator. When exposed to the "high" androstadienone concentration, women showed stronger hypothalamic activation than men. By contrast, men showed more hypothalamic activation when exposed to the "medium" androstadienone concentrations in comparison to women. Thus, we replicated that smelling the chemo-signal androstadienone elicits a hypothalamic activation. However, this effect does not seem to be gender-specific, because androstadienone activated the hypothalamus in both men and women, suggesting that androstadienone exerts specific effects in heterosexual individuals of both sexes. [less ▲]

The neural mechanisms controlling sexual behavior are sexually differentiated by the perinatal actions of sex steroid hormones. We recently observed using female mice deficient in alpha-fetoprotein (AFP-KO) and which lack the protective actions of AFP against maternal estradiol, that exposure to prenatal estradiol completely defeminized the potential to show lordosis behavior in adulthood. Furthermore, AFP-KO females failed to show any male-directed mate preferences following treatment with estradiol and progesterone, indicating a reduced sexual motivation to seek out the male. In the present study, we asked whether neural responses to male- and female-derived odors are also affected in AFP-KO female mice. Therefore, we compared patterns of Fos, the protein product of the immediate early gene, c-fos, commonly used as a marker of neuronal activation, between wild-type (WT) and AFP-KO female mice following exposure to male or estrous female urine. We also tested WT males to confirm the previously observed sex differences in neural responses to male urinary odors. Interestingly, AFP-KO females showed normal, female-like Fos responses, i.e. exposure to urinary odors from male but not estrous female mice induced equivalent levels of Fos protein in the accessory olfactory pathways (e.g. the medial part of the preoptic nucleus, the bed nucleus of the stria terminalis, the amygdala, and the lateral part of the ventromedial hypothalamic nucleus) as well as in the main olfactory pathways (e.g. the piriform cortex and the anterior cortical amygdaloid nucleus), as WT females. By contrast, WT males did not show any significant induction of Fos protein in these brain areas upon exposure to either male or estrous female urinary odors. These results thus suggest that prenatal estradiol is not involved in the sexual differentiation of neural Fos responses to male-derived odors. [less ▲]

We recently reported that female aromatase knockout (ArKO) mice show deficits in sexual behaviour and a decreased population of kisspeptin-immunoreactive neurones in the rostral periventricular area of ... [more ▼]

We recently reported that female aromatase knockout (ArKO) mice show deficits in sexual behaviour and a decreased population of kisspeptin-immunoreactive neurones in the rostral periventricular area of the third ventricle (RP3V), resurrecting the question of whether oestradiol actively contributes to female-typical sexual differentiation. To further address this question, we assessed the capacity of ArKO mice to generate a steroid-induced luteinising hormone (LH) surge. Adult, gonadectomised wild-type (WT) and ArKO mice were given silastic oestradiol implants s.c. and, 1 week later, received s.c. injections of either oestradiol benzoate (EB) followed by progesterone, EB alone, or no additional steroids to activate gonadotrophin-releasing hormone (GnRH) neurones and generate an LH surge. Treatment with EB and progesterone induced significant Fos/GnRH double-labelling and, consequently, an LH surge in female WT and in ArKO mice of both sexes but not in male WT mice. ArKO mice of both sexes had fewer cells expressing Kiss-1 mRNA in the RP3V compared to female WT mice but had more Kiss-1 mRNA-expressing cells compared to WT males, reflecting an incomplete sexual differentiation of this system. To determine the number of cells expressing kisspeptin, the same experimental design was repeated in Experiment 2 with the addition of groups of WT and ArKO mice that were given EB + progesterone and sacrificed 2 h before the expected LH surge. No differences were observed in the number of kisspeptin-immunoreactive cells 2 h before and at the time of the LH surge. The finding that ArKO mice of both sexes have a competent LH surge system suggests that oestradiol has predominantly defeminising actions on the GnRH/LH surge system in males and that the steroid-induced LH surge can occur in females even with a greatly reduced population of kisspeptin neurones in the RP3V. [less ▲]

We mapped cells immunoreactive for the phosphorylated form (p44/p42) of the mitogen-activated protein kinase (pMAPK--also known as ERK1/2) in the brain of male mice after exposure to female olfactory cues ... [more ▼]

We mapped cells immunoreactive for the phosphorylated form (p44/p42) of the mitogen-activated protein kinase (pMAPK--also known as ERK1/2) in the brain of male mice after exposure to female olfactory cues or after the display of male copulatory behaviors. Exposure to soiled bedding from estrous females or the display of coital behaviors rapidly (within 10 min) induced MAPK phosphorylation in most of the brain regions known to be involved in the processing of olfactory cues (main and accessory olfactory bulbs, amygdala, and medial preoptic area) and in the control of copulatory behavior (amygdala and medial preoptic area). MAPK phosphorylation thus seems to be a useful marker to study short-term neural activation associated with the expression of specific behaviors. [less ▲]